利用SSR标记定位东乡野生稻苗期耐冷性基因

应用由 2 13个株系组成的协青早B/东乡野生稻的BC1F1群体 ,分析了东乡野生稻的耐冷基因与DNA标记的连锁关系。以苗期的死苗率为指标 ,对亲本和BC1F1群体各株系进行耐冷鉴定。结果表明 :死苗率在BC1F1群体中呈连续分布 ,表明耐冷性是由多基因控制的数量性状。应用单因素方差分析法 ,分别在第 4、第8染色体上发现有与耐冷性连锁的SSR标记RM 2 80、RM 337。     

不结球白菜抗芜菁花叶病毒基因的遗传分析及SSR标记

为了找到与不结球白菜抗芜菁花叶病毒(TuMV)基因连锁的分子标记,采用群体分离分析法(BSA法)和SSR标记进行了与抗病基因连锁标记的筛选。通过对亲本、F1、BC1和F2群体进行病毒接种,研究了试验材料对TuMV的抗性遗传模型,结果表明:不结球白菜对TuMV的抗性由显性单基因控制。通过SSR引物筛选,得到在双亲间稳定表现多态性的引物54对。用这些引物筛选抗感池,得到1个与芜菁花叶病毒抗病基因连锁的SSR标记Ra3E05,连锁距离为8.7 cM。将该标记测序,得到1条184 bp的序列,根据该序列重新设计引物,将SSR标记转化为序列特异扩展区城(SCAR)标记SCRa2,用F2群体对其验证,带型与原SSR标记表现一致,可用于分子标记辅助育种。     

黄瓜白粉病抗性序列相关扩增多态性的分子标记研究

以高抗和高感白粉病的黄瓜亲本及其杂交后代F2分离群体为材料,应用序列相关扩增多态性(sequence-related amplified polymorphism,SRAP)技术开展与黄瓜白粉病抗性基因连锁的SRAP分子标记研究.结果表明:从437对SRAP引物组合中筛选出62对能够在2个亲本抗、感池间扩增出稳定多态性条带的引物组合,进一步用这些引物分析F2群体,发现有1对引物组合Mel/Em9扩增出的SRAP标记(Mel/Em9-284 bp)能与黄瓜抗白粉病基因连锁,遗传距离为9.8 cM.获得的分子标记对克隆抗病基因和利用分子标记辅助选择提高育种效率具有重要意义.     

与黄瓜抗黑星病基因连锁的分子标记研究

【目的】筛选与黄瓜抗黑星病基因连锁的分子标记，探讨黄瓜抗病材料鉴定和选择的分子方法。【方法】以黄瓜抗黑星病和感黑星病亲本组合（Q6×Q12）的F2分离群体为试材，采用BSA法和AFLP技术筛选与黄瓜抗黑星病基因连锁的分子标记。【结果】AFLP引物组合E20M64在抗病池和感病池间约130 bp处表现多态性。经F2单株分析，在抗病个体中扩增出了约为130 bp的特异片段，而感病个体无此条带。该特异标记与黑星病抗性基因紧密连锁，遗传距离为4.83 cM。测序结果显示，目标片段的大小为125 bp，并将该标记转换为了SCAR标记。【结论】提供了黄瓜黑星病抗性鉴定的分子手段，可用于分子标记辅助育种。     

与黄瓜全雌性基因连锁的SSR分子标记

黄瓜(Cucumis sativus L.)是重要的蔬菜栽培作物,其雌花率的高低直接影响着黄瓜产量。目前优良的黄瓜品种都具备全雌性或强雌性特征,全雌性也是黄瓜优势育种的重要途径。但由于黄瓜性别表现受到遗传和环境等多种因素的影响,传统的从表型上进行全雌性基因的选择效率不高。然而,借助与目的基因相连锁的分子标记进行辅助育种能直接从基因型上对后代单株进行选择,准确率高,能够在苗期进行性型鉴定,从而大大地提高育种效率。以全雌品系240-1-2-2-3-1自交系和弱雌品系3-5-1-3-2-1-1-1-1-2及其F1、F2、BC1P1和BC1P2世代为试验材料,进行田间鉴定和遗传规律分析。结果表明:黄瓜性别表达由寡基因控制,并受到一些背景基因的修饰;黄瓜全雌性相关基因遗传模型符合加性-显性-上位性遗传模型。利用PCR技术和SSR分子标记方法,通过亲本、F2全雌和全雄基因池筛选,从699对SSR引物组合中获得稳定的多态性引物组合2对,即CSWCT25和SSR18956;经回收、测序,特异片段全长分别为331bp和145bp,与黄瓜全雌性基因的连锁距离分别为7.7cM和6.8cM,均可用于黄瓜全雌系品种的辅助选育。     

甜瓜霜霉病抗性基因的SSR标记

为了研究与甜瓜抗霜霉病基因紧密连锁的SSR分子标记,以甜瓜抗病品种PI414723和感病品种DF4的杂交F2群体156个单株为试验材料,利用人工鉴定方法,根据F2抗、感病单株分离比例组建抗感池,用SSR技术寻找与抗病基因连锁的分子标记。从211个SSR引物组合中筛选出3个与抗病基因紧密连锁的标记DE1887、DE1320、DM0854,遗传距离分别为26、26、13.69 cM。     

SSR方法标记桃[Prunus persic(L.)Batsch]果肉近核色素(英文)

[目的]利用 SSR 分子标记法标记桃(Prunus persic (L.) Batsch)果肉近核色素。[方法]以"重阳红"与"燕红"2 个桃品种为亲本构建正交 F1群体,选取其中138株后代作为标记群体,采用分离群体分组分析(bulked segregate analysis,BSA)法,将果肉近核色素分为"有"和"无"2个基因池,应用SSR分子标记技术寻找与桃果肉近核色素性状基因连锁的分子标记。[结果]通过对256对引物的筛选,获得了3对与控制桃果肉近核色素性状基因连锁的分子标记,即 UDP96-003、ch04g09 和 UDP97-402,同时计算得到这 3 个标记与桃果肉近核色素性状基因的遗传距离分别为16.7、10.1和17.0 cM。[结论]该研究为进一步筛选遗传距离更近的共显性分子标记奠定了基础。     

SSR方法标记桃果肉近核色素

[目的]利用SSR分子标记法标记桃(Prunus persica(L.)Batsch)果肉近核色素。[方法]以"重阳红"与"金保"2个桃品种为亲本构建正交F1群体,选取其中138株后代作为标记群体,采用分离群体分组分析(bulked segregant analysis,BSA)法,将果肉近核色素分为"有"和"无"2个基因池,应用SSR分子标记技术寻找与桃果肉近核色素性状基因连锁的分子标记。[结果]通过对256对引物的筛选,获得了3对与控制桃果肉近核色素性状基因连锁的分子标记,即UDP96-003、ch04g09和UDP97-402,同时计算得到这3个标记与桃果肉近核色素性状基因的遗传距离分别为16.7、10.1和17.0 cM。[结论]该研究为进一步筛选遗传距离更近的共显性分子标记奠定了基础。     

水稻芽期耐冷性QTL的分子定位

 以籼粳交密阳23号/吉冷1号的F2：3 代200个家系作为作图群体，构建了一张含有97个微卫星 （SSR）标记的分子连锁图谱。在5℃低温条件下，对F3家系进行芽期耐冷性鉴定，并利用SSR标记进行了芽期耐冷性数量性状位点（QTL）分析。研究结果表明，芽期耐冷性在F3家系群中呈单峰连续分布，表现为由多基因控制的数量性状；共检测到与芽期耐冷性有关的QTL 3个，分别位于第2、4 和7染色体上，对表型变异的贡献率范围为11.5%～20.5%。其中，位于第4染色体RM273～RM303的qCTBP4对表型变异的贡献率最大。     

美洲南瓜(Cucurbit pepo)WMV-2抗性遗传及分子标记研究

以美洲南瓜(Cucurbita pepo)抗病自交系‘0504’和感病自交系‘0223’为亲本杂交获得的157个F2单株为材料,研究南瓜对西瓜花叶病毒2号(WMV-2)抗性的遗传规律和分子标记。对亲本、F1及F2苗期接种WMV-2进行抗病性鉴定,结果表明,美洲南瓜对WMV-2的抗性由单隐性基因控制。采用集团分离分析法(BSA)和SSR技术,筛选出与WMV-2抗性基因连锁的标记CMTm157和CMTm158,遗传距离分别为10.9cM和6.2cM,其可以作为南瓜抗WMV-2育种的辅助选择分子标记。     

黄瓜白粉病抗性基因定位及候选基因分析

【目的】通过对黄瓜白粉病抗性基因的精细定位,明确抗性基因的候选区段,为进一步克隆基因及解析基因功能奠定基础。【方法】本研究选用高抗白粉病资源(自交系‘74’)和感白粉病资源(自交系‘80’)配制杂交组合,构建F_1、F_2群体,利用单囊壳白粉菌鉴定亲本及群体白粉病抗性并进行遗传分析;采用BSA-seq的方法对黄瓜白粉病抗性基因进行初步定位,在此基础上在候选区段开发分子标记,进一步精细定位白粉病抗性基因。【结果】分离群体单株表型鉴定结果表明,自交系74的抗性是由不完全隐性基因控制的。BSA-seq技术初步将抗性基因定位于5号染色体15—25 Mb位置,命名为PM 74。最终,利用分子标记将抗性基因定位于SSR15321和SSR07531之间,遗传距离为3.06 c M,物理距离为238 444 bp,可解释41.95%的表型变异。生物信息学分析表明在候选区段内包含有17个候选基因,其中Cucsa.275630为TIR-NBS-LRR类基因。【结论】本研究将黄瓜白粉病抗性基因精细定位到约238 kb的候选区段内,区段内包含有一个TIR-NBS-LRR基因,该基因将是今后研究的重点候选基因。     

Inheritance Analysis and Identification of SSR Markers Linked to Late Blight Resistant Gene in Tomato

Late blight caused by Phytophthora infestans is the most serious disease of tomato production in China. Studies on the genetics of resistance and identification of molecular markers are very useful for breeding late blight resistant varieties. The objective of this paper was to study the inheritance of late blight resistance and identify simple sequence repeat （SSR） markers associated with resistance allele in tomato （Lycopersicon esculentum Mill）. The results came from an F2 progeny of 241 plants derived from a cross between 5~ inbred line that is susceptible to late blight and a resistant accession CLN2037E. The late blight responses of F2 plants were tested by artificially inoculation of detached-leaflets in plate and natural infection assayed under greenhouse conditions. Both methods showed that the resistance is dominant and inherited as monogenic trait. Genetic mapping and linkage analysis showed that the late blight resistance gene Ph-ROL was located on chromosome 9 with a genetic distance of 5.7 cM to the SSR marker TOM236.     

Inheritance and Molecular Mapping of Stripe Rust Resistance Gene Yr88375 in Chinese Wheat Line Zhongliang 88375

Stripe rust is one of the most important diseases of wheat worldwide. Inheritance of stripe rust resistance and mapping of resistance gene with simple sequence repeat （SSR） markers are studied to formulate efficient strategies for breeding cultivars resistant to stripe rust. Zhongliang 88375, a common wheat line, is highly resistant to all three rusts of wheat in China. The gene conferring rust disease was deduced originating from Elytrigia intermedium. Genetic analysis of Zhongliang 88375 indicated that the resistance to PST race CYR31 was controlled by a single dominant gene, temporarily designated as Yr88375. To molecular map Yr88375, a F2 segregating population consisting of 163 individuals was constructed on the basis of the hybridization between Zhongliang 88375 and a susceptible wheat line Mingxian 169; 320 SSR primer pairs were used for analyzing the genetic linkage relation. Six SSR markers, Xgwm335, Xwmc289, Xwmc810, Xgdmll6, Xbarc59, and Xwmc783, are linked to Yr88375 as they were all located on chromosome 5BL Yr88375 was also located on that chromosome arm, closely linked to Xgdmll6 and Xwmc810 with genetic distances of 3.1 and 3.9 cM, respectively. The furthest marker Xwmc783 was 13.5 cM to Yr88375. Hence, pedigree analysis of Zhongliang 88375 combined with SSR markers supports the conclusion that the highly resistance gene Yr88375 derived from Elytrigia intermedium is a novel gene for resistance to stripe rust in wheat. It could play an important role in wheat breeding programs for stripe rust resistance.     

Molecular Mapping of Two Novel Stripe Rust Resistant Genes YrTp1 and YrTp2 in A-3 Derived from Triticum aestivum × Thinopyrum ponticum

Loss of variety resistance to stripe rust (Puccinia striiformis Westend f. sp.tritici) is an important factor causing massive periodical epidemic of rust in wheat production. Creation and development of new races of rust pathogen have led to serious crisis of resistance loss in widely planted varieties. This has quickened the search for new resistance resources. Molecular marker could facilitate the identification of the location of novel genes. A line A-3 with high resistance (immune) to currently epidemic yellow rust races (CY29, 31, 32) was screened out in offspring of Triticum aestivum × Thinopyrum ponticum. Segregation in F₂ and BC₁ populations indicated that the resistance was controlled by two independent genes: one dominant and one recessive. SSR markers were employed to map the two resistant genes in the F₂ and BC₁ populations. A marker WMC477-_167bp located on 2BS was linked to the dominant gene with genetic distance of 0.4 cM. Another marker WMC364-_{208 bp} located on 7BS was linked to the recessive-resistant gene with genetic distance of 5.8 cM. The two genes identified in this paper might be two novel stripe rust resistant genes, which were temporarily designated as YrTp1 and YrTp2, respectively. The tightly linking markers facilitate transfer of the two resistant genes into the new varieties to control epidemic of yellow rust.     

Genetic analysis and SSR mapping of stem rust resistance gene from wheat mutant D51

Wheat (Triticum aestivum L.) stem rust caused by Puccinia graminis f. sp. tritici is one of the main diseases of wheat worldwide. Wheat mutant line D51, which forms a highly susceptive cultivar ‘L6239’ to the three races notated and cultured with immature embryos, shows resistance to prevailing races 21C3CPH, 21C3CKH, and 21C3CTR of P. graminis f. sp. tritici in China. In this study, the number and the expression stages of the resistance genes in mutant D51 were studied using inoculation identification and microsatellite (SSR) marker analysis. Two F₁ populations from the crosses of D51 × L6239 (60 individuals) and D51 × Chinese Spring (60 individuals), their F₂ populations (185 and 175 individuals respectively) at the seedling stage, and one F₂ population derived from the cross of D51 × L6239 (194 individuals) at the adult stage were inoculated with pathogen race 21C3CPH to test for resistance. All F₁ individuals of the two crosses were immune to stem rust at both seedling and adult stages. The response pattern of the three F₂ populations showed that the R:S segregation ratio was 3:1, suggesting that the stem rust resistance of D51 is controlled by a single dominant gene, and is expressed during the entire growth period. The identification of the stem rust resistance by the F₃ progeny test confirmed the credibility of the F₂ population test. Segregating populations and small population analyses were used to identify chromosomal regions and molecular markers linked to the gene by the SSR marker method. A total of 675 SSR markers and 185 individuals of the D516L6239 F2 population were used to search genetically linked markers to the target gene. Using Mapmaker 3.0 and Map-draw with Kosambi’s function and other options set at default values, molecular mapping revealed that the gene was located on chromosome 5DS, linked with and flanked by two SSR markers, Xgwm190 and Xwmc150, at 18.58 and 21.33 cM, respectively. It has been reported that only one stem rust resistant gene, Sr30, is located on the wheat chromosome 5DL, and that it has no resistance to 34C2MKK and 34C2MFK, while the parent L6239 of mutant D51 has no resistance to 21C3CPH, 21C3CTK and 21C3CTR, but has resistance to 34C2MKK and 34C2MFK. The results above indicate that the gene identified in the study might be a novel resistance gene to stem rust, tentatively designated as SrD51. __________ Translated from Acta Agronomica Sinica, 2007, 33(8): 1262–1266 [译自: 作物学报]     

Genetic Analysis and Preliminary Mapping of a Highly Male-Sterile Gene in Foxtail Millet (Setaria italica L. Beauv.) Using SSR Markers

Breeding of male-sterile lines has become the mainstream for the heterosis utilization in foxtail millet, but the genetic basis of most male-sterile lines used for the hybrid is still an area to be elucidated. In this study, a highly male-sterile line Gao146A was investigated. Genetic analysis indicated that the highly male-sterile phenotype was controlled by a single recessive gene a single recessive gene. Using F₂ population derived from cross Gao146A/K103, one gene controlling the highly male-sterility, tentatively named as ms1, which linked to SSR marker b234 with genetic distance of 16.7 cM, was mapped on the chromosome VI. These results not only laid the foundation for fine mapping of this highly male-sterile gene, but also helped to accelerate the improvement of highly male-sterile lines by using molecular marker assisted breeding method.     

Inheritance and Molecular Mapping of Stripe Rust Resistance Gene Yr88375 in Chinese Wheat Line Zhongliang 88375

Stripe rust is one of the most important diseases of wheat worldwide. Inheritance of stripe rust resistance and mapping of resistance gene with simple sequence repeat (SSR) markers are studied to formulate efficient strategies for breeding cultivars resistant to stripe rust. Zhongliang 88375, a common wheat line, is highly resistant to all three rusts of wheat in China. The gene conferring rust disease was deduced originating from Elytrigia intermedium. Genetic analysis of Zhongliang 88375 indicated that the resistance to PST race CYR31 was controlled by a single dominant gene, temporarily designated as Yr88375. To molecular map Yr88375, a F2 segregating population consisting of 163 individuals was constructed on the basis of the hybridization between Zhongliang 88375 and a susceptible wheat line Mingxian 169; 320 SSR primer pairs were used for analyzing the genetic linkage relation. Six SSR markers, Xgwm335, Xwmc289, Xwmc810, Xgdm116, Xbarc59, and Xwmc783, are linked to Yr88375 as they were all located on chromosome 5BL. Yr88375 was also located on that chromosome arm, closely linked to Xgdmll6 and Xwmc810 with genetic distances of 3.1 and 3.9 cM, respectively. The furthest marker Xwmc783 was 13.5 cM to Yr88375. Hence, pedigree analysis of Zhongliang 88375 combined with SSR markers supports the conclusion that the highly resistance gene Yr88375 derived from Elytrigia intermedium is a novel gene for resistance to stripe rust in wheat. It could play an important role in wheat breeding programs for stripe rust resistance.     

Identification and Molecular Mapping of the RsDmR Locus Conferring Resistance to Downy Mildew at Seedling Stage in Radish (Raphanus sativus L.)

Downy mildew (DM), caused by the fungus Peronospora parasitica, is a destructive disease of radish (Raphanus sativus L.) worldwide. Host resistance has been considered as an attractive and environmentally friendly approach to control the disease. However, the genetic mechanisms of resistance in radish to the pathogen remain unknown. To determine the inheritance of resistance to DM, F₁, F₂ and BC₁F₁ populations derived from reciprocal crosses between a resistant line NAU-dhp08 and a susceptible line NAU-qtbjq-06 were evaluated for their responses to DM at seedling stage. All F₁ hybrid plants showed high resistance to DM and maternal effect was not detected. The segregation for resistant to susceptible individuals statistically fitted a 3:1 ratio in two F₂ populations (F_2(SR) and F_2(RS)), and 1:1 ratio in two BC₁F₁ populations, indicating that resistance to DM at seedling stage in radish was controlled by a single dominant locus designated as RsDmR. A total of 1972 primer pairs (1036 SRAP, 628 RAPD, 126 RGA, 110 EST-SSR and 72 ISSR) were screened, and 36 were polymorphic between the resistant and susceptible bulks, and consequently used for genotyping individuals in the F₂ population. Three markers (Em9/ga24₃₇₀, NAUISSR826₇₀₀ and Me7/em10₄₀₀) linked to the RsDmR locus within a 10.0 cM distance were identified using bulked segregant analysis (BSA). The SRAP marker Em9/ga24₃₇₀ was the most tightly linked one with a distance of 2.3 cM to RsDmR. These markers tightly linked to the RsDmR locus would facilitate marker-assisted selection and resistance gene pyramiding in radish breeding programs.     

黄瓜ZYMV抗性鉴定与抗性基因的分子标记辅助选择

近年来,小西葫芦黄化花叶病毒(zucchini yellow mosaic virus,ZYMV)在黄瓜(Cucumis sativus L.)上危害愈发严重,影响黄瓜植株的生长发育,降低黄瓜的产量与品质。由于ZYMV缺乏有效的防治药剂,因此需要培育ZYMV抗性黄瓜品种。以ZYMV Z5-1为病毒源,对不同黄瓜材料的ZYMV抗性进行鉴定,并对抗性位点候选基因进行了测序分析。结果显示,黄瓜品种SA02、SA06、S94、S1003、WI7230与TMG-1具有ZYMV抗性,其抗性候选基因Csa6G152960.1的突变位点一致。在ZYMV抗性鉴定的基础上,通过回交转育,将黄瓜自交系TMG-1中的ZYMV抗性基因转入感病自交系SA0422中,回交期间利用分子标记对回交群体进行前景选择和背景选择,经过3代回交,将ZYMV抗性基因转入SA0422,为培育抗ZYMV黄瓜新品种奠定了基础。     

分子标记辅助甜瓜抗蔓枯病基因的聚合及品种改良

【目的】蔓枯病（gummy stem blight）是一种真菌性土传病害,是危害甜瓜生产的严重病害之一,其存在生理小种的分化,品种即使仅携带单个抗病基因,仍会导致甜瓜抗性逐步降低甚至丢失。建立甜瓜抗蔓枯病聚合育种的分子标记辅助选择体系,选育抗性高且不再分离的聚合抗源自交系和品质优良且高抗甜瓜蔓枯病的改良白皮脆品种（品系）,为甜瓜优质、抗病和高产育种提供一种简单、快捷的选择方法和重要的中间材料。【方法】利用5份甜瓜单一抗源材料PI140471、PI157082、PI511890、PI482398和PI420145两两杂交获得8份聚合抗源（082-471、082-890、082-398、145-471、145-082、145-890、145-398和890-398）。利用3个不同浓度的蔓枯病病菌孢子液（5×10⁵个/mL、5×10⁷个/mL、5×10⁹个/mL）对8份聚合抗源的F₁植株幼苗进行接种鉴定。结合春、秋两季的接种鉴定结果,筛选得到2份聚合后抗性显著提高的基因组合145-471和145-398。一方面,以145-471和145-398为材料,利用苗期梯度接种鉴定、分子标记辅助选择以及农艺性状观察,对其自交后代进行逐代筛选至F₇。另一方面,选用组合PI420145和PI140471以及PI420145和PI482398作为蔓枯病抗性基因的供体亲本,综合性状优良的感病甜瓜品种‘白皮脆’为抗性基因受体亲本。先分别单向回交5代,在单个基因回交转移时,利用分子标记检测结合农艺性状观察对回交后代进行筛选。当回交后代的遗传背景得到一定程度恢复后,再将2个方向的回交后代进行杂交,后自交3代稳定。【结果】聚合抗源对不同浓度的蔓枯病菌均表现为抗,而单一抗源对不同浓度蔓枯病菌表现出选择性抗性且抗性水平低于聚合抗源。SSR标记CMCT505和CMTA170a在PI140471和PI482398上可分别扩增出189 bp和121 bp的特异性片段,SCAR标记SGSB₁₈₀₀可以在PI420145上扩增出1 800 bp的特异性片段,而145-471（145-398）的聚合单株可以同时扩增出189 bp和1 800 bp（121 bp和1 800 bp）两条特异片段。聚合抗源145-471（或145-398）的自交F₇世代以及改良白皮脆BC₅F₄世代群体的分子标记鉴定结果显示,各单株已经成功聚合了Gsb-1和Gsb-6（或Gsb-4和Gsb-6）两个抗病基因,抗性不再有分离且农艺性状稳定一致。初步建立了甜瓜抗蔓枯病聚合育种的分子标记辅助选择体系,获得了两份表现高抗且抗性不再分离的聚合抗源自交系F₇和2个以优质品种‘白皮脆’为受体亲本的改良白皮脆BC₅F₄品种（系）。改良白皮脆BC₅F₄世代表现为高抗甜瓜蔓枯病,且在单果质量、果形指数、果实脆度、果肉质地、果肉厚度和可溶性固形物含量等农艺性状方面与白皮脆并无显著差异。【结论】创建的3个分子标记CMCT505、CMTA170a和SGSB₁₈₀₀对抗病基因Gsb-1、Gsb-4和Gsb-6的选择具有较高的准确性。初步建立了甜瓜抗蔓枯病聚合育种的分子标记辅助选择体系,为甜瓜优质、抗病和高产育种提供了一种简单、快捷的选择方法,将大大提高育种的效率。改良‘白皮脆’作为抗病育种的新材料,为甜瓜抗病品种的选育和抗病基因进一步聚合提供了材料。